dsRNA binding properties of RDE-4 and TRBP reflect their distinct roles in RNAi.
Double-stranded RNA (dsRNA)-binding proteins facilitate Dicer functions in RNA interference. Caenorhabditis elegans RDE-4 facilitates cleavage of long dsRNA to small interfering RNA (siRNA), while human trans-activation response RNA-binding protein (TRBP) functions downstream to pass siRNA to the RNA-induced silencing complex. We show that these distinct in vivo roles are reflected in in vitro binding properties. RDE-4 preferentially binds long dsRNA, while TRBP binds siRNA with an affinity that is independent of dsRNA length. These properties are mechanistically based on the fact that RDE-4 binds cooperatively, via contributions from multiple domains, while TRBP binds noncooperatively. Our studies offer a paradigm for how dsRNA-binding proteins, which are not sequence specific, discern dsRNA length. Additionally, analyses of the ability of RDE-4 deletion constructs and RDE-4/TRBP chimeras to reconstitute Dicer activity suggest RDE-4 promotes activity using its dsRNA-binding motif 2 to bind dsRNA, its linker region to interact with Dicer, and its C-terminus for Dicer activation.
Parker
Greg S
GS
University of Utah, Department of Biochemistry/HHMI, Salt Lake City, UT 84112-5650, USA.
Maity
Tuhin Subhra
TS
Bass
Brenda L
BL
D000818:Animals; D017173:Caenorhabditis elegans / Q000502:physiology*; D029742:Caenorhabditis elegans Proteins / Q000378:metabolism*; D006801:Humans; D011485:Protein Binding; D034622:RNA Interference; D012330:RNA, Double-Stranded / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism*; D000099253:Trans-Activation Responsive RNA-Binding Protein
D029742:Caenorhabditis elegans Proteins; D012330:RNA, Double-Stranded; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; D000099253:Trans-Activation Responsive RNA-Binding Protein; C461645:RDE-4 protein, C elegans